ABSTRACTS
Previous EHRS Annual Meetings ...2
Welcome ... 3
General Information ...5
Accommodation Information ...5
Social Events ...5
The Programme at a Glance...6
Scientific Programme...7 Invited Lectures...17 Oral Communications...23 Poster Presentations...79 Proceedings ...141 …about Delphi ...147
The International Anthem of the EHRS...153
2
(Greece), Janssen-Cilag (Greece), Novartis (Greece), Pfizer (Greece), Sanofi-Aventis (Greece), Servier (Greece), Astellas (Greece), AstraZeneca (Greece), Roche (Greece), Vianex (Greece), Wyeth (Greece), Novo Nordisk (Denmark), UCB (Belgium), Abbott (USA), Hoffmann-LaRoche (Switzerland), Chembiotin (Greece)
Poster Design: Dimitrios Kourkoutis
Previous EHRS Annual Meetings
1972 Paris; 1973 Marburg; 1974 Copenhagen; 1975 Florence; 1976 Paris; 1977 London; 1978 Lodz; 1979 Stockholm; 1980 Visegrád; 1981 Hannover; 1982 Bled; 1983 Brighton; 1984 Florence; 1985 Aachen; 1986 Odense; 1987 Strbske Pleso; 1988 Copenhagen; 1989 Breda; 1990 Kuopio; 1991 Marburg; 1992 Malaga; 1993 Cologne; 1994 Budapest; 1995 Moscow; 1996 Antwerp; 1997 Seville; 1998 Lodz; 1999 Lyon; 2000 Nemi (Rome); 2001 Turku; 2002 Eger; 2003 Noordwijkerhout; 2004 Cologne; 2005 Bled.
3 10 May 2006
Myth and science meet at Delphi, the navel (Gk: omphalos) of the earth that owes its
reputation to the oracle of Apollo, where the god's divination was given through the mouth of his prophetess Pythia.
On behalf of the Organising Committee, I welcome you to the XXXVth Meeting of the European Histamine Research Society at the European Cultural Centre of Delphi. This annual EHRS Meeting is organised, for the first time in Greece, by the Department of Pharmacology of the Medical School of the University of Athens.
The Organizers and the Scientific Committee made every effort to ensure that all
presentations are filled with the scents and the sounds of the most motivating up-to-date scientific information bound together in a radiant circle of histamine research. The sessions will focus on the chemistry and pharmacology of the four histamine receptor subtypes, on established and novel functions of the amine in inflammation, cognition, metabolism, reproduction, differentiation, immunology, neuro-immuno-endocrinology, cancer and genomics. Three stimulating invited talks will surround these topics. The president of the EHRS, Prof M. Ennis will guide the audience through the achievements in histamine research in the 35 year long history of the Society; Prof A. Falus will introduce the future perspectives of histamine in systems biology; Prof P. Maestrelli will portray the clinical dimension in the “D Varonos Memorial Symposium”. The continued interest in histamine research will be revealed in the competitive “Art Hancock Young Investigator Award Symposium”. Finally, we are delighted that Ingrid Uvnäs will be presented with an Honorary Membership, for her support to our Society over the years.
Thanks are due to the participants who made this event happen. Travelling across
paths paved with unlimited affection for research, they came from their homelands to this corner of the Mediterranean to disclose effective roles of histamine in an effort to ameliorate the art of life. This journey would be impossible without the valuable assistance of our PhD students and the kind support of the sponsors, who contributed to the materialization of this histamine feast around the oracle of Delphi, under the spirit of wondering principles of our ancestors.
Delphi is a place of inexhaustible mythological and historic references. The motivating
scientific sessions together with the archaeological site of Delphi, the picturesque varied scenery of Mount Parnassos, the olive tree valley, the Corinthian Gulf harbours of Galaxidi, Kirra and Itea and the contemporary cultural life and natural beauty of the region hopefully will provide infinite opportunities for a most pleasant stay.
Welcome to Delphi!
On behalf of the organising committee
Catherine Tiligada
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Location of the Meeting
European Cultural Centre of Delphi (ECCD), Greece
Host Institute
Department of Experimental Pharmacology
Medical School, University of Athens, M. Asias 75, GR-11527 Athens, Greece
President of the EHRS
Professor Madeleine Ennis
The Queen's University of Belfast, UK
Chairperson of the Meeting
Dr Ekaterini Tiligada
University of Athens Medical School, Greece
Organising Committee Andreas Delitheos Madeleine Ennis Vassiliki Giannoulaki Sotirios Kakavas Zoe Papadopoulou-Daifoti Christina Spyraki Ekaterini Tiligada Executive Assistants Vassilios Delitheos Konstantinos Papamichael Evangelia Zampeli
Abstract Evaluation & Bursary Selection Committee
Madeleine Ennis (Chairperson) Patrizio Blandina
Andras Falus
Tatjana Irman Florjanc Pertti Panula
Gill Sturman Anita Sydbom
The Art Hancock Young Investigator Award Jury
Walter Schunack (Chairman), Timothy A Esbenshade, Pertti Panula
Poster Jury
Frank Ahrens (Chairman), El-Sayed K Assem, Patrizio Blandina, Bernhard F Gibbs, Nina Grosman, Mariusz Gujski, Krisztina Hegyi, Zsuzsanna Huszti,
Tatjana Irman-Florjanc, Emanuela Masini, Danuta Maślińska, Elena S Rivera, Elke Schneider, Holger Stark, Gill Sturman, Anita Sydbom
5 Wednesday May 10th 15:00 – 21:00 at the Delphi Palace Hotel
Thursday May 11th 08.00 – 13.00 at the ECCD Friday May 12th 08.00 – 18.00 at the ECCD Saturday May 13th 08.00 – 13.00 at the ECCD
Congress Website and E-Mail / Internet Service
http://conferences.med.uoa.gr/ehrs-06
E-mail and Internet services is provided at the Delphi Palace Hotel
Oral Communications
They should last no longer than 10 min (plus 5 min for discussion), exept for the EUBREN Basophil Network Symposium, which is scheduled for 15 min (plus 5 min discussion). A PC equipped with MS PowerPoint 2003 for Data Projection is available. The presentations should be brought in a USB memory stick (MS PowerPoint format). Presentation preview: On Wednesday May 10th, 15.00-21.00, at the Registration desk in Delphi Palace Hotel and the following days during the first coffee break, in the ECCD Conference Hall.
Poster Presentations
The posters will be on display throughout the meeting. Presenters should present the main point(s) from the poster in 1 min (followed by 4 min discussion) in the corresponding session. They are encouraged to stand by their posters during the coffee breaks. Short-listing posters for the poster prizes (announced on Saturday 13th, 14.00) will be revisited by the Poster Jury on Saturday 13th, 16.30-17.00. Prizes will be announced at the Farewell Dinner.
Accommodation Information
Guest House “EUROPA”
Delphi Palace 69 Apollonos Str, 33054 Delphi Tel: +30 2265082151 Hotel King Iniohos 78 Osiou Louka Str, 33054 Delphi Tel: +30 2265082701 Hotel Iniohos 19 Vas. Pavlou & Friderikis, 33054 Delphi Tel: +30 2265082710
Social Events
Wednesday May 10th 19:30 Welcome reception on the terrace of “Delphi Palace Hotel” (dress: informal)
Thursday May 11th 15:00 Visit to the Delphi archaeological site and museum
17:30 Visit to the Monastery of “Profitis Elias” 19:00 Visit and dinner in the harbor of Galaxidi.
(dress: mid-season clothes, comfortable shoes, light sweatshirt/raincoat)
Friday May 12th 20:00 Dinner at the Piano Restaurant “Iniohos”
(dress: informal)
Saturday May 13th 20:00 Farewell Dinner at the “Villa Symposium” (dress: lounge suit/elegant dress recommended but not essential)
Transfer to Athens International Airport “El. Venizelos” on Sunday 14th
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18.00 - 19.30 EHRS Council Meeting 19.30 - 22.30 Welcome reception
Thursday 11 May 2006
08.00 - 13.00 Registration 08.00 - 08.45 Opening ceremony
08.45 - 09.30 Invited lecture: “Histamine and the EHRS – what has happened in the
last 35 years?” Madeleine Ennis
09.30 - 10.30 Oral session 1: Future perspectives in histamine research
10.30 - 11.45 Coffee break
10.45 - 11.45 Poster session 1: H3 receptors
Poster session 2: Mast cells and Basophils
11.45 - 13.00 Oral session 2: Allergy and Inflammation
13.00 - 13.30 Poster session 3: Histamine in the CNS Poster session 4: Histamine in the GI tract
13.30 - 14.30 Lunch
15.00 - 22.00 Excursion - Dinner
Friday 12 May 2006
08.00 - 19.00 Registration Desk Open
08.00 - 08.45 GB WEST lecture: “Histamine and systems biology; genes and genomics
beyond genes” Andras Falus
08.45 - 09.45 Oral session 3: Metabolism, Reproduction and Cancer
09.45 - 11.15 Oral session 4: Histamine and brain
11.15 - 12.00 Coffee break
11.30 - 12.00 Poster session 5: Clinical aspects of histamine research Poster session 6: Histamine in Reproduction and Cancer
12.00 - 13.30 The Art Hancock Young Investigator Award Symposium
13.30 - 14.30 Lunch
14.30 - 17.00 The Art Hancock Memorial Symposium
17.00 - 18.00 Coffee break
17.15 - 17.50 Poster session 7: Immunology
Poster session 8: Receptors and other subjects
20.15 - 22.00 Dinner
Saturday 13 May 2006
08.30 - 13.00 Registration Desk Open
08.30 - 09.15 Invited lecture: “Occupational asthma: diagnosis and management”
Piero Maestrelli
09.15 - 10.30 The D Varonos Memorial Symposium: “Clinical implications – histamine
and disease” 10.30 - 11.00 Coffee break
11.00 - 12.50 Oral session 5: Neuro-immuno-endocrine aspects of histamine
13.00 - 14.00 Lunch
14.00 - 16.30 The EUBREN Basophil network symposium
16.30 - 17.00 Coffee break
16.30 - 17.00 Poster evaluation Committee 17.15 - 19.00 General assembly
20.30 FarewellDinner
Sunday 14 May 2006
7 after the farewell dinner in the early hours of Sunday 14 May 2006.
Wednesday 10 May 2006
15.00-21.00 Arrival - Check in - Registration 18.00-19.30 EHRS Council Meeting 19.30-22.30 Welcome reception
Thursday 11 May 2006
08.00-13.00 Registration08.00-08.45 Opening ceremony
08.45-09.30 I1: Invited lecture
Histamine and the EHRS – what has happened in the last 35 years?
Madeleine Ennis, UK
Introduced by Prof Fred L Pearce
09.30-10.30 Oral session 1: Future perspectives in histamine research
Chairpersons: Prof Fred Pearce & Prof Agnieszka Fogel 09.30-09.45 O1: Novel ligands stabilize stereo-selective conformations of the H1
receptor to result in functionally-selective activation or antagonism of intracellular signaling pathways
Booth RG
09.45-10.00 O2: Fast production and analysis of the human histamine H1 receptors using a cell free protein expression system followed by MS analysis
Sansuk K, Bakker RA, Hensbergen P, Leurs R
10.00-10.15 O3: Effect of sulphasalazine and balsalazide on histamine release from mast cells
Peh KH, Wan BCY, Assem ESK, Pearce FL
10.15-10.30 O4: Luminometric determination of amine oxidase activity Schwelberger HG, Feurle J
10.30-11.45 Coffee break
10.45-11.45 Poster session 1: H3 receptors
Chairpersons: Prof Patrizio Blandina & Prof Holger Stark P1: H3 receptor activation phosphorylates Akt in rat cortical neurons
Mariottini C, Chiarugi A, Fossati S, Blandina P, Passani MB P2: Histamine H3 receptors modulate [3H]-dopamine release in rat
substantia nigra pars reticulata, but not in prefrontal cortex Ramos-Jiménez J, Garduño-Torres B, Camacho J, Arias-Montaño J-A
P3: Is the high affinity selective H3 receptor agonist methimepip anxiolytic or anxiogenic in a new validated animal models of human anxiety?
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of immethridine, a potent and highly selective histamine H3-R agonist
Celanire S, Gillard M, Christophe B, Quere L, Collart P, Dassesse D, van Marle A, Hulscher S, Bakker R, Zuiderveld P, Timmerman H, Leurs R, Chatelain P, Lamberty Y, Talaga P P6: Histamine H3 receptors regulate glutamate, but not GABA
release in rat thalamus
Garduño-Torres B, Treviño M, Gutiérrez R, Arias-Montaño J-A P7: Heterocyclic replacements for the central phenyl-core of
diamine–based human histamine H3 receptor antagonists Carruthers NI
P8: Histamine 3 receptor inverse agonists for the treatment of obesity. Biological and chemical challenges
Freichel C, Gatti-Mac Arthur S, Hertel C, Huwyler J, Nakagawa T, Nettekoven M, Plancher J-M, Raab S, Richter H, Roche O, Sarmiento RMR, Schuler F, Taylor S, Ullmer C, Wiegand R P9: Ureas with H3-antagonist activity–a new scaffold discovered by
lead-hopping starting from cinnamic acid amides Lau J, Jensen CB, Lund P, Hohlweg R
P10: Radiosynthesis and biodistribution of a histamine H3 receptor antagonist: evaluation of a potential pet ligand
Airaksinen AJ, Jablonowski JA, Van der May M, Barbier AJ, Klok RP, Verbeek J, Schuit R, Herscheid JDM, Leysen JE, Carruthers NI, Lammertsma AA, Windhorst AD
P11: (3-Piperidin-1-yl-propoxy)-tetrahydroisoquinolines and tetra-hydroazepines: A novel series of selective histamine H3 receptor antagonists
Jesudason CD, Beavers LS, Cramer JW, Dill J, Finley DR, Gleason SD, Hemrick-Luecke SK, Lindsley CW, Nelson DLG, Stevens FC, Gadski RA, Oldham SW, Pickard RT, Siedem CS, Sindelar DK, Singh A, Watson BM, Witkin JM, Hipskind PA
10.45-11.45 Poster session 2: Mast cells and Basophils
Chairpersons: Dr Nina Grosman & Dr El-Sayed K Assem P12: Mast cell-derived interleukin-8 may be involved in the ovarian
mechanisms of follicle growth and ovulation
Szukiewicz D, Pyzlak M, Klimkiewicz J, Szewczyk G, Maślińska D
P13: Potent histamine-releasing activity of atrahagin, a snake venom metalloproteinase
Wei JF, Mo YZ, Qiao LY, Wei XL, Chen HQ, Xie H, Fu YL, He SH
P14: Gene expression profiling of mouse mucosal mast cell (mMMC) proteases
9 Maśliński S
P16: Clinico-biological characteristics of flow cytometry applied to NSAIDs hypersensitivity diagnosis. Determination of preliminary ROC curves
Sainte-Laudy J, Touraine F, Boumediene A, Cogné M
P17: Amitriptyline inhibits mast cell histamine secretion: implications for chronic fatigue syndrome therapy
Theoharides TC, Kempuraj D
P18: Lack of endogenous histamine affects early inflammatory phase of wound healing
Holub MC, Falus A
P19: Transtympanic versus intramuscular steroid administration in a histamine-induced inflammatory middle ear model
Chimona TS, Panayiotides JG, Papadakis CE, Helidonis ES, Velegrakis GA
P20: A heretofore undisclosed crux of eosinophilia-myalgia syndrome: compromised histamine degradation
Smith MJ, Garrett RH
P21: Music therapy, “adverse” diet and histamine
Hanke A, Klawitter B, Meng H, Herwald M, Hasenmaier M, Borck H, Fischer M, Diel E, Diel F
P22: Analysis of human basophil activation feed back induced by histamine and high histamine dilutions
Sainte-Laudy J, Belon Ph
P23: Whithdrawn
11.45-13.00 Oral session 2: Allergy and Inflammation
Chairpersons: Prof Friedhelm Diel & Prof Marija Čarman-Kržan 11.45-12.00 O5: Activation of cannabinoid receptors prevents allergen-induced
asthma-like reaction in sensitized guinea pigs
Fabrizi F, Vannacci A, Giannini L, Mariottini C, Passani MB, Mannaioni PF, Nistri S, Masini E
12.00-12.15 O6: Effects of the carbon monoxide releasing molecule CORM-3 in a coincubation model with rat mast cells and human neutrophils Vannacci A, Giannini L, Fabrizi F, Uliva C, Mastroianni R, Masini E, Mannaioni PF
12.15-12.30 O7: Prevention of antigen-induced bronchoconstriction by epigallocatechine-3-gallate in actively sensitized guinea pigs Giannini L, Uliva C, Bani D, Mastroianni R, Suzuki Y, Nistri S, Mannaioni PF, Suzuki H, Masini E
12.30-12.45 O8: Increased nitric oxide production mediates T cell signal transduction in histidine decarboxylase knockout mice
Koncz A, Nagy G, Mazan M, Buzás E, Falus A
12.45-13.00 O9: Histamine influences the DNA interaction of STATs in human lymphocytes
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P25: Excitatory effect of histamine on neuronal activity of rat globus pallidus by activation of H2 receptors in vitro
Chen K, Wang J-J
P26: Whithdrawn
P27: Histamine improves rat rota-rod and balance beam performances through H2 receptors in the cerebellar interpositus nucleus
Song Y-N, Li H-Z, Zhu J-N, Wang J-J
P28: P2Y receptor mediated excitation in the posterior hypothalamus Sergeeva OA, Klyuch BP, Fleischer W, Eriksson K, Korotkova TM, Siebler M, Haas HL
P29: Central histaminergic system – the regulatory mechanism of circulatory homeostasis in haemorrhagic shock
Jochem J
13.00-13.30 Poster session 4: Histamine in the GI tract
Chairpersons: Dr Frank Ahrens & Dr Hegyi Krisztina
P30 Amitriptyline affects guinea pig post-heparin plasma diamine oxidase activity
Rajtar S, Irman-Florjanc T
P31: Immunolocalization of histamine H3 receptor in the rat gastrointestinal tract
Chazot PL, Shenton FC, Schunack W, Grandi D, Morini G P32: Influence of H3/H4 receptor antagonist thioperamide on regional
haemodynamics in rats with trinitrobenzene sulfonic acid-induced colitis
Fogel AW, Jochem J, Lewinski A
P33: Whithdrawn
P34: Functional gut mucosal biopsy testing using tolerated food antigens during mucosa oxygenation yields a high extent of specificity in non-atopic, non-allergic healthy individuals Raithel M, Sehnert C, Nägel A, Backhaus B, Straube S, Buchwald W, Schultis W, Kressel J, Hahn EG, Kimpel S, Konturek P
P35: Mechanisms underlying the radioprotective effect of histamine on small intestine
Medina VA, Mohamad NA, Croci M, Cricco GP, Núñez MA, Martín GA, Bergoc RM,Rivera ES
13.30-14.30 Lunch
11 08.00-19.00 Registration Desk Open
08.00-08.45 I2: GB WEST lecture
Histamine and systems biology; genes and genomics beyond genes
Andras Falus, Hungary
Introduced by Prof Madeleine Ennis
08.45-09.45 Oral session 3: Metabolism, Reproduction and Cancer
Chairpersons: Prof Madeleine Ennis & Dr Hubert Schwelberger 08.45-09.00 O10: Amine system project
Medina MA, Aldana JF, Villatoro FR, Claros G, Urdiales JL, Trelles O, Sánchez-Jiménez F
09.00-09.15 O11: Effects of histamine on triglyceride breakdown in mouse adipocytes
Carpéné C, Bour S, Prévot D, Duffaut C, Valet P 09.15-09.30 O12: Highlights in histamine function and reproduction
Pap E, Pállinger E, Falus A
09.30-09.45 O13: Expression of histamine H3 and H4 receptors in benign lesions and malignant carcinomas of the human mammary gland Medina VA, Croci M, Crescenti EJV, Bergoc RM, Rivera ES
09.45-11.15 Oral session 4: Histamine and brain
Chairpersons: Dr Paul Chazot & Dr Zoe Papadopoulou-Daifoti 09.45-10.00 O14: Generation of the first anti-hH4 receptor antibody: identification
of the H4R in human lymphocytes and brain
Chazot PL, Shenton FC, Van Rijn RM, Bakker RA, Leurs R 10.00-10.15 O15: Aversive memory consolidation and reconsolidation: differences
in neurotransmitter engagement.
Bucherelli C, Baldi E, Mariottini C, Passani MB, Blandina P 10.15-10.30 O16: Properties of native GABAA receptors in histaminergic and
orexinergic neurons Sergeeva OA, Haas HL
10.30-10.45 O17: Characteristics of cortical EEG and sleep-wake cycle in histamine H1-receptor knockout mice
Parmentier R, Anaclet C, Watanabe T, Lin J-S
10.45-11.00 O18: Histamine and behavioral state-dependent control of hippocampal plasticity
Selbach O, Haas HL 11.00-11.15 O19: Whithdrawn
11.15 -12.00 Coffee break
11.30-12.00 Poster session 5: Clinical aspects of histamine research
Chairpersons: Dr Anita Sydbom & Dr Mariusz Gujski P36: Whithdrawn
P37: Evaluation of urinary N-methylhistamine excretion during a long-term follow up of patients with inactive Crohn`s disease Kimpel S, Nägel A, Backhaus B, Straube S, Buchwald W, Schultis W, Kressel J, Hahn EG, Raithel M
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P39: Do both systemic and topical anti-allergy treatments cause ocular dryness?
Gomes P,Abelson MB, Torkildsen G
P40: Use of rEV131 for prevention of anterior chamber inflammation in an animal model
Gomes P, Lane K, Abelson MB, Weston-Davies W
P41: Lidocaine protein binding time response to diphenhydramine administration in rat tissues.
Tigka E, Daskala I, Tsagatakis M, Melakopoulos I, Saranteas Th, Tesseromatis C
11.30-12.00 Poster session 6: Histamine in Reproduction and Cancer
Chairpersons: Prof Elena S Rivera & Prof Danuta Maślińska P42: Locally secreted histamine may regulate the development of
ovarian follicles by apoptosis
Szukiewicz D, Klimkiewicz J, Pyzlak M, Szewczyk G, Maślińska D
P43: Histamine H4 receptors in human placenta in diabetes-complicated pregnancy.
Szewczyk G, Maślińska D, Szukiewicz D, Deregowski K, Smiertka W, Klimkiewicz J, Pyzlak M
P44: Histamine increase the invasive potential of human melanoma cells
Pocza P, Kovacs P, Pallinger E, Pocza K, Kohidai L, Falus A, Darvas Z
P45: Endogenous and exogenous histamine affects on growth of mice mammary adenocarcinoma
Hegyesi H, Molnár V, Fulop KA, Falus A
P46: Nitric oxide involvement in histamine-mediated PANC-1 cells growth
Cricco GrP, Medina VA, Núñez MA, Mohamad NA, Gutiérrez AS, Sanbuco LA, Bergoc RM, Rivera ES, Martín GA
P47: Differential modulation of the cellular stress response by histamine and the H1, H2 receptor antagonists in eukaryotic cells Delitheos V, Papamichael K, Tiligada E
12.00-13.30 The Art Hancock Young Investigator Award Symposium
Chairpersons: Prof Walter Schunack & Prof Pertti Panula 12.00-12.15 O20: Oligomerization of the human histamine H4 receptor
van Rijn RM, Chazot PL, Shenton FC, van Marle A, Bakker RA, Leurs R
12.15-12.30 O21: Homology modeling of the human histamine H4 receptor and its application in structure-based drug design
Kiss R, Jelinek I, Falus A, Noszál B
12.30-12.45 O22: Characterization of 4-methylhistamine and VUF 8430 as selective histamine H4R agonists
Lim HD, Guaita E, Bakker RA, Coruzzi G, Thurmond RL, Leurs R
13 13.00-13.15 O24: Enhanced CREB activity and adipogenic potential in histamine
free embryonic fibroblasts Hegyi K, Falus A, Toth S
13.15-13.30 O25: Excitation of aminergic neurons by histamine
Korotkova TM, Sergeeva OA, Ponomarenko AA, Haas HL 13.30-14.30 Lunch
14.30-17.00 The Art Hancock Memorial Symposium
Chairpersons: Dr Timothy A Esbenshade & Prof Helmut L Haas 14.30-14.45 Art Hancock: the man, his achievements and his life at the
EHRS
Timothy A Esbenshade
14.45-15.00 O26: Biochemical and pharmacological evidence for homo- and heter-oligomeric human H3 receptor isoforms: further evidence of dominant negative role for splice isoforms
Chazot PL, Shenton FC, van Rijn RM, Bakker RA, LeursR 15.00-15.15 O27: New signaling pathways for the histamine H3 receptor
Bongers G, van Marle A, Navis M, Bakker RA, Leurs R
15.15-15.30 O28: Histamine H3 receptor expression is increased in the brains of transgenic mice over-expressing beta amyloid
Ning X, Liu D, Pong K, Pangalos M, Reinhart P, Hirst W 15.30-15.45 O29: Different response to GABAA or H3 antagonists suggest the
existence of distinct subpopulations among histaminegic neurons
Giannoni P, Cenni G, Passani MB, Mannaioni PF, Medhurst AD, BlandinaP
15.45-16.00 O30: Cloning and expression of histamine H1, H2 and H3 receptors in zebrafish and effects of their ligands on behavior
Panula P, Peitsaro N, Sundvik M
16.00-16.15 O31: A new family of histamine H3 receptor antagonists based on a natural product: discovery, development of SAR, and properties of the series
Cowart M, Sun M, Zhao C, Witte DG, Miller TR, Krueger KM, Browman K, Fox GB, Esbenshade TA, Hancock AA
16.15-16.30 O32: Histamine H3 antagonists with serotonin reuptake transporter inhibitor activity
Letavic MA
16.30-16.45 O33: Pharmacological classification of histamine H3 receptor agents across species is attributable to TM3 sequence differences Gibbs BF, Estvander BR, Miller TR, Baranowski JL, Sharma R, Hancock AA, Krueger KM
16.45-17.00 O34: Fluorophore-tagged non-imidazole histamine H3 receptor ligands with subnanomolar affinities
Amon M, Ligneau X, Schwartz JC, Stark H 17.00-18.00 Coffee break
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P49: Histamine H1, H2 but not H4 receptors are upregulated during bone marrow regeneration
Horváth Z, Pállinger E, Horváth G, Jelinek I, Falus A
P50: Unexpected action of histamine on dendritic cells under acidosis Amaral MM, Davio C, Martínez D, Geffner J, Vermeulen M P51: Tissue-related immune-endocrine interactions in experimental
hyperthyroidism
Kakavas S, Zampeli E, Delitheos V, Tiligada E
P52: Histamine stimulates electrogenic ion transport in avian epithelia
Collins CB, Campion DP, McGrath J, Baird AW
P53: Peripheral but not central effects of allergic reaction are mediated by histamine
Gerencsér AS, Kovács KJ
P54: Study of adipogenesis in genetically histamine free (HDC KO) and wild type bone marrow derived mesenchymal stem cells Lux A, Skard I, Toth SP, Falus A
17.15-17.50 Poster session 8: Receptors and other subjects
Chairpersons: Prof Tatjana Irman-Florjanc & Prof Emanuela Masini
P55: Internalization and resensitization mechanisms of the histamine type 2 receptor
Fernández N, Monczor F, Notcovich C, Baldi A, Davio C, Shayo C
P56: Strain specific differences in Oct4 expression of HDC KO and wild type ES cells
Toth SP, Gocza E, Carstea BV, Falus A
P57: Comparison of the in vitro and in vivo H2-H4 receptor selectivity of 4-methylhistamine and VUF8430
Adami M, Guaita E, Lim HD, Bakker RA, de Esch IJP, Leurs R, Coruzzi G
P58: Porcine plasma amine oxidase has a broad substrate specificity and efficiently converts histamine
Feurle J, Schwelberger HG
P59: Influence of amitriptyline on central histamine-induced reversal of haemorrhagic shock in rats
Jochem J, Irman-Florjanc T, Zwirska-Korczala K
P60: Altered inflammatory gene expression in genetically H4 histamine receptor deficient mice
Fulop AK, Ribita D, Falus A
P61: Involvement of the histaminergic system in the central cardiovascular regulation in haemorrhage-shocked rats with portocaval anastomosis; preliminary data
Jochem J, Fogel AW, Maksymowicz M, Zwirska-Korczala K, Lewinski A
15 08.30-13.00 Registration Desk Open
08.30-09.15 I3: Invited lecture
Occupational asthma: diagnosis and management Piero Maestrelli, Italy
Introduced by Prof Pier Francesco Mannaioni
09.15-10.30 The D Varonos Memorial Symposium
“Clinical implications – histamine and disease”
Chairpersons: Prof Pier Francesco Mannaioni & Dr Catherine Tiligada
Sponsored by GlaxoSmithKline (Greece)
09.15-09.30 O35: The histamine H4 receptor mediates allergic airway inflammation Thurmond RL
09.30-09.45 O36: Distribution pattern of histamine H4 receptor in human synovial tissue from patients with rheumatoid arthritis
Jablonowska M, Grzybowska-Kowalczyk A, Wojtecka-Lukasik E, Maślińska D, Gujski M, Maśliński S
09.45-10.00 O37: Theophylline as “add-on” therapy to cetirizine in patients with chronic idiopathic urticaria: a randomized, double-blind, placebo-controlled pilot study
Makris M, Kalogeromitros D, Kempuraj D, Katsarou-Katsari A, Grigoriou S, Theoharides TC
10.00-10.15 O38: Biphasic immunomodulating effect of cepharolsporin derivatives in man in vitro and in vivo
Assem ESK, Ezeamuzie CI, Vickers MR
10.15-10.30 O39: On the mechanism of antiphagocyte-antioxidative effect of H1-antihistamines
Nosal R, Drabikova K, Jancinova V, Pecivova J, Macickova T, Holomanova D
10.30-11.00 Coffee break
11.00-12.50 Oral session 5: Neuro-immuno-endocrine aspects of
histamine
Chairpersons: Prof Wilfried Lorenz & Dr Robin Thurmond 11.00-11.35 O40: Evidence of human mast cell function in inflammatory diseases
Theoharides TC,Cao J, Papadopoulou N,Kempuraj D, Tagen M 11.35-11.50 O41: Whithdrawn
11.50-12.05 O42: Skin histidine decarboxylase (HDC) and corticotropin-releasing hormone receptor-1 (CRH-R1) genes are overexpressed in chronic urticaria (CU)
Kalogeromitros D, Papadopoulou N, Staurianeas NG, Tiblalexi D, Theoharides TC
12.05-12.20 O43: Role of histamine in ghrelin-induced gastroprotection against acute gastric lesions
Konturek PC, Brzozowski T, Pajdo R, Konturek SJ, HahnEG, Raithel M
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12.35-12.50 O45: G protein receptor kinases (GPKs) regulate histamine H2R desensitization in human leukemic cells
Fernández N, Monczor F, Tubio MR, Baldi A, Shayo C, Davio C 13.00-14.00 Lunch
14.00-16.30 The EUBREN Basophil network symposium
Chairpersons: Dr Franco Falcone & Prof Clemens Dahinden 14.00-14.10 Introduction to EUBREN
Franco Falcone
14.10-14.30 O46: Control of basophil functions through histamine and other biogenic amines
Schneider E, Machavoine F, Pléau J-M, Ohtsu H, Watanabe T, Schinkel AH, Dy M
14.30-14.50 O47: In vitro and in-vivo stimulatory potential of bone marrow-derived mast cell on tumor growth
Hegyesi H, Darvas Z, Wiener Z, Molnár V, Pos Z, Falus A 14.50-15.10 O48: Substantial differences in the kinetics of histamine release from
human basophils caused by varying strengths of IgE-dependent activators
Gibbs BF,Räthling A, Zillikens D, Huber M, Haas H
15.10-15.30 O49: The IL-4-inducing principle from S. mansoni eggs (IPSE) activates human basophils via a novel mechanism: “IgE receptor engagement without crosslinking”
Blindow S, Schramm G, Gronow A, Manske M, Galle J, Grevelding CG, Weimar T, Sutton BJ, Gibbs BF, Doenhoff MJ, Haas H
15.30-15.50 O50: Accumulation and activation of basophils in allergic disease Walls AF, Mochizuki A, McEuen AR
15.50-16.10 O51: Old and novel mediators of Th2-type immune responses and allergy produced upon mast cell – basophil interactions
Dahinden CA, Spiegl N, Tschopp CM, Didichenko S
16.10-16.30 O52: “British diet on a chip”: A novel diagnostic tool for the detection of food allergy combining protein microarrays with human basophils
Lin J, Schramm G, Haas H, Falcone FH, Alcocer MJC 16.30-17.00 Coffee break
16.30-17.00 Poster evaluation Committee
17.15-19.00 General assembly
20.30 Farewell Dinner
Sunday 14 May 2006
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18
Histamine and the EHRS - What Has Happened in the Last 35 Years? Madeleine Ennis
Respiratory Research Group, The Queen’s University of Belfast, Belfast BT12 6BJ, UK Histamine has a long history compared to many other mediators. In 1907 it was synthesized by Windaus and Vogt and in 1910 it was discovered in nature (Ackermann and Dale & Barger). The idea of a histamine society began in 1971, at a histamine satellite meeting in Lodz organised by the late Czeslaw Maslinski (one of our few Honorary Members). The first official meeting of the then named “Histamine Club” took place in Paris in 1972. This was the same year as Sir James Black (another of our honorary members) and colleagues described the discovery of the H2 receptor. In the intervening years since that very first meeting, the number of known histamine receptors has increased to four. We now realise that histamine also acts as a modulator of the immune system and is involved in angiogenesis, cancer as well as the allergic diseases. In this talk I will try and highlight some of the important presentations from our meetings showing how the histamine field has developed. One of the characteristics of our society is its friendliness and the fact that we have enough time for informal contacts. These are really important and often lead to some fantastic scientific collaborations. We also learn something about the countries we visit (so far 16) during our outings and again have the opportunity to network with others.
In the intervening years, we have moved from the Histamine Club to the European Histamine Research Society – complete with a registered address in Belgium and statutes in French as well as members from such “well-known European Countries” such as Argentina, Canada, China, Japan, Mexico and USA (nb this is not an exhaustive list). Last year, Professor Takehiko Watanabe became our first non-European Honorary Member of the Society.
19 Andras Falus
Department of Genetics, Cell and Immunobiology, Semmelweis University, Nagyvarad ter 4, 1089 Budapest, Hungary
Systems biology is the study of living organisms in terms of their underlying network structure rather than simply their individual molecular components. A "system" can be anything from a gene regulatory network to a cell, a tissue, or an entire organism. Because systems biology requires investigation of all interacting components simultaneously, high-throughput, quantitative technologies are essential. Computational approaches are also required to handle and interpret the volumes of data necessary to understand complex biological systems.
Histamine biology, pharmacology and medicine provide eminent examples for the need for multilateral approaches. This colorful scenario is traditionally represented in sessions of European Histamine Research Society. SNP and SNP haplotype studies, gene expression and proteomic analysis are currently complemented with metabolomic approaches as well as computational analysis from allergy databanks, since computational models are important complementary methodologies to experimental research. They are particularly useful in fields requiring a large number of experiments due to the combinatorial nature of the underlying systems and processes. In such fields, computational models evolve with the amount of data and the accumulation of knowledge.
Further and even more challenging perspectives are seen on the field of non-coding DNA (PostGenes) or as called earlier the „junk DNA”. It turned out that there is a forty times larger difference between the DNA of the chimp and the human in the 98.7% "junkDNA" as compared to 1.3% encoding for proteins. Although the relevance is largely unclear, one of the functions of the PostGene part of genome is coding microRNA hairpins with outstanding, newly recognized importance in gene regulation. In histamine genomics there are fairly good indications, that further attention will be paid to the non-coding PostGenes providing new haploidomics predictive and thus experimentally supportable or refutable theories in histaminology.
20
Occupational Asthma: Diagnosis and Management Piero Maestrelli
Department of Environmental Medicine and Public Health, University of Padova, Padova, Italy Introduction
Occupational asthma (OA) is defined as a type of asthma that is caused by exposure to a product present in the workplace. To fulfill this definition, the causative agent(s) should be almost exclusive to the workplace, hence this definition excludes asthma triggered by physical agents such as cold air or exercise. Besides OA with a latency period for which an allergic mechanism can be identified or is highly probable, another form of work-related airflow obstruction and bronchial hyperresponsiveness has also been described, i.e. irritant-induced asthma or reactive airway dysfunction syndrome (RADS), which may be developed after acute exposure to high concentrations of irritant gases or fumes. RADS therefore develops without a latency period of exposure and, although its functional characteristics resemble those of OA, its symptoms cannot be reproduced by re-exposure of the affected patients to nonirritant amounts of the offending agent.
OA may be induced by several different mechanisms. Immunological mechanisms are generally implicated when OA occurs upon exposure to an agent after a latent period of sensitization. These mechanisms can be further divided into those that induce asthma through either an IgE-dependent or an apparently non-IgE-dependent mechanism. In the latter, specific IgE antibodies are not detectable or are found only in a small proportion of the patients with proven disease, even though the clinical picture is compatible with an ‘allergic’ reaction (i.e. sensitization and an exaggerated tissue damaging response upon re-exposure). Nonimmunological mechanisms are implicated when OA occurs without a latency period of exposure, i.e. RADS or irritant-induced asthma.
The causes of OA can be classified into high- and low-molecular-weight compounds. High-molecular-weight compounds, which are often from biological sources, generally induce asthma through an IgE-dependent mechanism, whereas the majority of low-molecular-weight compounds induce asthma through non-IgE-dependent mechanisms.
Clinical history
Since an attributable risk of asthma due to occupation of 15% among all asthma cases has been estimated, every subject with asthma should be questioned about his or her current and past workplaces as persistent asthma can be attributed to past exposure. Two clues in the patient’s history should point to the possibility of OA: the symptoms, and the job and products at work. First, the subject may have a clear-cut history of exacerbations of respiratory symptoms at work. Secondly, the job and product at work can suggest the diagnosis. The nature of all products present in the workplace, not only those handled by the subject, should be obtained by requesting safety data sheets. There could be products present that have not been listed as known causes of OA, but this does not preclude the possibility of there being so. At the onset of symptoms, improvement at weekends and when on vacation is generally the rule, although eventually the symptoms will persist through these short periods away from work.. It must be remembered that questionnaires are sensitive, but that they are not specific tools, when results are compared with the final diagnosis.
21 presence of immediate skin reactivity or increased specific IgE or IgG may reflect exposure or sensitization but it does not imply that the target organ is involved. With high-molecular-weight allergens, negative skin tests to such allergens almost completely exclude the possibility of OA. With low-molecular-weight allergens, such as isocyanates and red cedar, negative skin tests or specific IgE or IgG do not refute or confirm the diagnosis of OA; skin tests are also usually unavailable.
Specific antibodies to allergens may be demonstrated in biological fluids using a variety of tests. They confirm a sensitization demonstrated by skin test but are often less sensitive. They represent an alternative to the skin test when the preparations of antigens have irritant, toxic, or mutagenic effects, and in patients under pharmacological treatment that blunts normal skin reactivity. Control for specificity is required, especially when protein conjugates are used. Different factors, such as total IgE level, characteristics of the conjugate, carrier specificity, and cross-reactivity with other antigens, may affect the results. Assessment of the chemokine monocyte chemoattractant protein-1 (MCP-1) produced in vitro by diisocyanate-stimulated blood mononuclear cells exhibited higher test efficiency than specific antibodies for identification of isocyanate asthma. Other in vitro tests, such as histamine release from basophils, are less standardized but may be useful occasionally. Physiological assessment
The presence of airway obstruction with demonstrable reversibility after inhaling a bronchodilator is a well-recognized confirmatory step for asthma. If there is no significant airway obstruction, the demonstration of increased bronchial responsiveness is suggestive of asthma, not necessarily of OA.
Serial PEF monitoring has been proposed for both the investigation and assessment of asthma. The sensitivity and specificity of PEF monitoring, as compared with the ‘gold standard’ specific inhalation challenges, varies from 72% to 89% depending on the study. Combining PEF and the assessment of eosinophils in induced sputum for periods at work and away from work may improve the diagnostic yield.
Laboratory challenges in small cubicles were proposed by Pepys in the 1970s in an effort to reproduce the workplace environment. Improvement in the methodology of the test has been put forward using closed-circuit apparatus for dry particles and vapors, including isocyanates. After exposure, various temporal patterns of reactions can occur, including those of typical (immediate, late, and dual) and atypical reactions (progressive, square waved, and prolonged immediate).
Outcome
The outcome of OA after diagnosis is often poor. Removal from the occupational exposure is associated with recovery from asthma in about 50% of subjects. Many retrospective studies have unanimously demonstrated a persistence of asthmatic symptoms, bronchial obstruction, and hyperresponsiveness in subjects with OA after being removed from exposure. Most studies also showed that the total duration of exposure, the duration of exposure after the onset of symptoms, and the severity of the asthma at the time of diagnosis are all determinants of the prognosis. Improvement in bronchial responsiveness occurs predominantly in the first two years after cessation of exposure and continues, though at a slower rate, later on. If exposure continues, there is overall deterioration in the asthmatic condition.
Mapp CE, Boschetto P, Maestrelli P, Fabbri LM. Am J Respir Crit Care Med 2005;172:280-305
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24
Novel Ligands Stabilize Stereo-Selective Conformations of the H1
Receptor to Result in Functionally-Selective Activation or Antagonism of Intracellular Signaling Pathways
Raymond G Booth
Department of Medicinal Chemistry, University of Florida, Gainesville, FL 32610-0485, USA Depending on its conformation, the histamine H1 GPCR can couple to several different G protein types to activate multiple intracellular signaling pathways. We developed novel ligands that stabilize stereo-selective conformations of the H1 receptor to result in functional selectivity. The cyclic tertiary amines, (±)-cis-5-phenyl-7-dimethylamino-5,6,7,8-tetrahydro-9H-benzocycloheptane (cis-PAB) and (–)-trans-1-phenyl-3-dimethylamino-1,2,3,4-tetrahydronaphthalene (trans-PAT), are functionally-selective agonists at H1 receptors, stimulating phospholipase (PL) C/inositol phosphate (IP) and adenylyl cyclase (AC)/cAMP signaling, respectively, in clonal cells and mammalian tissues. Cis-PAB and trans-PAT also are functionally-selective H1 antagonists of AC/cAMP and PLC/IP signaling, respectively. Molecular determinants for cis-PAB vs. trans-PAT conformationally-selective binding to H1 receptors, that leads to functionally-selective effects on PLC/IP vs. AC/cAMP signaling, likely involves stereochemical factors, as well as, more subtle steric influences. In bovine adrenal cells, tyrosine hydroxylase (TH) and catecholamine synthesis is activated (EC50 +/- SEM, µM; Emax +/- SEM at 10 µM, % control) by histamine (0.16 +/- 0.08; 200+/- 4), cis-PAB (1.0 +/- 0.1; 160 +/- 14), and trans-PAT (1.4 +/- 0.1; 160 +/- 5.3) in a manner blocked by H1 antagonists. Inhibition of PKA, but, not PKC, abolishes the histamine and trans-PAT effect, suggesting H1 agonism by these ligands exclusively activates AC/cAMP/PKA signaling to stimulate TH. Inhibition of PKA or PKC abolishes the cis-PAB effect. Since PAB activation of H1 receptors selectively stimulates PLC/IP signaling (antagonizes AC/cAMP signaling), it appears H1/PLC/IP/PKC signaling can activate PKA, downstream of cAMP production, to stimulate TH. Thus, there appears to be PKC to PKA signaling “cross-talk” that can confound prediction of functional selectivity effects downstream of second messenger formation. Support: NIMH 068655.
25
1
Using a Cell Free Protein Expression System Followed by MS Analysis Kamonchanok Sansuk1, Remko Bakker1, Paul Hensbergen, Rob Leurs1
1Leiden/Amsterdam Center for Drug Research (LACDR), Vrije Universiteit Amsterdam,
Department of Pharmacochemistry, De Boelelaan 1083, 1081 HV, Amsterdam; 2Biomolecular
Mass Spectrometry Unit, Leiden University Medical Center (LUMC), Department of Parasitology, P.O. box 9600, 2300 RC, Leiden, The Netherlands
Rational design of ligands for G protein coupled receptors (GPCRs) relies for a large extent on receptor models. Computer models of GPCRs, however, are commonly based on bovine rhodopsin for which a high resolution 3D GPCR structure is available, in conjunction with site-directed mutagenesis data for the GPCR at hand. In order to produce better models structural data of the studied receptor would be needed. One of the requirements for structural studies is to have access to large amounts of pure GPCR protein.
Using a baculovirus expression system we previously successfully produced large amounts of the human histamine H1 receptor (hH1R) (Ratnala VRP et al., Eur. J. Biochem. 2004; 271:2636-2646). Here we have employed a cell free expression system to produce large quantities of the hH1R. We have tried to optimize solubilization, purification and reconstitution of the produced hH1R for subsequent structural analysis. Using MALDI-TOF (Matrix Assisted Laser Desorption Ionization-Time Of Flight) mass spectrometry we were able to detect a large percentage of the hH1R consisting of both intra- and extracellular loops, amino- and carboxyl-termini as well as transmembrane domains. The use of a cell free expression system enables a much faster production of protein compared to baculovirus expression, whereas MS analysis requires much smaller amounts of protein for analysis compared to solid state NMR. In summary, the cell free expression combined with MS structural analysis enables a fast and specific screening of the target receptor.
26
Effect of Sulphasalazine and Balsalazide on Histamine Release from Mast Cells
Kheng H Peh1, Beatrice YC Wan1, El-Sayed K Assem2, Frederick L Pearce1
1Department of Chemistry, University College London, 20 Gordon Street, London WC1H 0AJ; 2Department of Pharmacology, University College London, Gower Street, London WC1E 6BT,
UK
Sulphasalazine (SAZ), balsalazide (BSZ) and their active moiety, 5-aminosalicyclic acid (ASA) offer effective treatment of ulcerative colitis. SAZ, BSZ and ASA inhibit histamine release (HR), the proinflammatory mediator, from mast cells (MC) [1,2]. We have studied the effect of SAZ, BSZ and ASA on HR from RBL-2H3 cells (mucosal type MC) and rat peritoneal MC (RPMC, connective tissue type MC).
RBL-2H3 cells [3] were pre-incubated with 5, 50 and 500 µM of test compounds for 30 min, followed by the addition of submaximal concentration of Con A (concanavalin A, 50 µg/ml, immunological stimulant), NaF (20 mM, non-specific G-protein activator), PLC (0.08 U/ml, phospholipase C) or A23187 (1.0 µM, calcium ionophore) for 15 or 30 min. For experiments with RPMC, cells from naive or Nippostrongylus brasiliensis-sensitised Sprague-Dawley rats (250-350 g) were preincubated with test compounds (0.16–500 µM) for 0 or 5 min prior to the challenge with anti-IgE serum (1/250 dilution), Con A (4 µg/ml), NaF (20 mM), PLC (0.08 U/ml), A23187 (10 µM) or DiC8 (PKC activator, 20 µM) for 10, 15 or 60 min [3].
With RBL cells, only NaF-induced HR was reduced by SAZ (51%, p<0.01) and BSZ (32%, p<0.05) at 500 µM but not ASA. No other induced HR was inhibited. Inhibition of Con A-induced HR by H2O2 (1 mM) was not reversed by SAZ, BSZ or ASA.
With RPMC, anti-IgE-induced HR was inhibited slightly by SAZ (at 20 µM, 12%, p<0.01) and markedly by BSZ (at 20µM, 48%, p<0.01; at 100 µM, 39%, p<0.05) but potentiated slightly by ASA (at 100–500 µM, 10–17%, p<0.05). Marked and dose-related inhibition of Con A-induced HR was shown by SAZ (at 20–500 µM, 41–79%, p<0.05, or <0.01) and BSZ (at 4–500 µM, 51–77%, p<0.01). NaF-induced HR was only inhibited by SAZ (24%, p<0.001) and ASA (14%, p<0.01). No other induced HR was inhibited.
With the exception of part of the results with NaF (RPMC), inhibition of induced HR by SAZ BSZ and ASA in the two cell preparations seems to be due to modulation of G-protein-controlled transmembrane signaling via diverse pathways.
[1] Peh KH et al. Br J Pharmacol 1990;100:449P [2] Bissonnette EY et al. J Immunol, 1996;156:218-23 [3] Wan BYC et al. Biochem Pharmacol 2001;62:1537-44
27 Hubert G Schwelberger, Johannes Feurle
Labor für Theoretische Chirurgie, Universitätsklinik für Chirurgie, Medizinische Universität Innsbruck, Austria
Amine oxidases play a key role in the inactivation of histamine. Diamine oxidase (DAO), a secreted, soluble copper-containing amine oxidase, catalyzes the direct oxidative deamination of histamine probably in the extracellular space. FAD-containing monoamine oxidases (MAO) of the outer mitochondrial membrane have been implicated in the oxidation of methylated histamine produced by histamine N-methyltransferase inside cells. Although numerous assays have been described for measurement of amine oxidase activity these either suffer the limitation of low sensitivity or require the use of certain amine substrates. Therefore, a new type of assay was developed that is not only extremely sensitive but also allows the measurement of the conversion of any amine substrate by any amine oxidase. In this assay, hydrogen peroxide, the common product of all amine oxidation reactions, is used by horseradish peroxidase to oxidize luminol in the presence of enhancers, which produces light that can be measured in a luminometer. This assay allows the simple assessment of substrate specificity, determination of kinetic parameters, study of enzyme inhibition mechanisms and investigation of long-term enzyme stability. The assay can be performed in single tube or microplate format and is very cost-efficient due to negligible reagent consumption.
28
Activation of Cannabinoid Receptors Prevents Allergen-Induced Asthma-Like Reaction in Sensitized Guinea Pigs
Francesca Fabrizi1, Alfredo Vannacci1,Lucia Giannini1, Chiara Mariottini1, Beatrice Passani1, Pier Francesco Mannaioni1, Silvia Nistri2, Emanuela Masini1
1Department of Preclinical and Clinical Pharmacology; 2Department of Anatomy, Histology and
Forensic Medicine, University of Florence, 50139 Florence, Italy
Asthma is defined by airway inflammation and hyperresponsiveness, and its morbidity has increased worldwide. The bronchodilating properties of Cannabis Indica derivatives are well known and recently the antinflammatory activity of CB2-receptor agonists in isolated guinea pig mast cells has been shown.
This study evaluates the effect of a synthetic CB receptor agonist (CP55,940) on asthma-like reactions to inhaled antigen in actively sensitized guinea pigs in vivo. Male albino guinea-pigs, sensitized to ovalbumin, placed in whole body respiratory chambers, were challenged with ovalbumin. CP55,940 (0.4 mg/Kg b.w.) were given i.p. 3 hours before ovalbumin challenge. Some animals, 30 min before CP55,940 treatment, received i.p. a CB1 (AM251, 0.1 mg/Kg b.w.) or a CB2 –receptor antagonist (SR144528, 0.1 mg/kg b.w.).
The following functional parameters were examined: latency time for the onset of respiratory abnormalities, cough severity score, occurrence and duration of dyspnoea. We also evaluated lung tissue histopathology, mast cell degranulation, lung tissue concentration of myeloperoxidase, malonyldialdehyde and 8-hydroxy-2-deoxyguanosine, as well as manganese superoxide dismutase and prostaglandin D2 in the bronchoalveolar lavage fluid (BAL).
Both respiratory abnormalities and bronchoconstriction in response to ovalbumin challenge are nearly absent in naïve animals, while they sharply became severe in sensitized animals. Treatment with CP55 significantly reduced the severity of cough and the occurrence of dyspnoea, and delayed the onset of respiratory abnormalities. Furthermore, all the biochemical changes induced by ovalbumin in lung tissue or BAL fluid were prevented by CP55 treatment. The pre-treatment of sensitized animals with CB1 (AM251) and CB2 (SR144528) –receptor antagonists reverted the protective effects of CP55, indicating that both CB1 and CB2 receptors are involved.
These results show that the cannabinoid receptor agonist CP55 can counteract acute allergic asthma-like reaction in actively sensitized guinea-pigs and suggest a possible future use of cannabinoid agonists in asthma.
29 Coincubation Model with Rat Mast Cells and Human Neutrophils
Alfredo Vannacci, Lucia Giannini, Francesca Fabrizi, Caterina Uliva, Rosanna Mastroianni, Emanuela Masini, Pier Francesco Mannaioni
Department of Preclinical and Clinical Pharmacology, University of Florence, viale Pieraccini 6, 50139, Florence, Italy
The heme oxygenase (HO) enzymes are able to split the tetrapyrrole heme ring to biliverdin, free ferrous iron, and carbon monoxide (CO). At least two isoforms of heme oxygenase are expressed in mammalian cells: HO-1 (inducible isoform) and HO-2 (constitutive). In particular, HO-1 is a stress-responsive enzyme that acts during inflammatory reactions, regulating immunological responses involved in cardiac anaphylaxis, in allergic reactions and in the rejection of transplanted organs. Previous reports from our group showed that exogenous CO or water-unsoluble CO-releasing molecules were able to mimic the anti-allergic and anti-anaphylactic effects of HO-1 in isolated guinea pig hearts, in guinea pig mast cells and in human basophils, mainly through the activation of the soluble guanylyl cyclase. Here we report the effects of the water soluble CO-releasing molecule CORM-3 in a coincubation of rat serosal mast cells (MCs) and human neutrophils (PMN), activated with the chemotactic peptide formyl-methionyl-leucyl-phenylalanine (fMLP, 10 nM). The expression of CD203c upon MC membrane (evaluated through flow cytometry) as well histamine release (fluorimetric method) were increased after the incubation with PMN stimulated by fMLP. CORM-3 (100 nM-10 uM) was able to reduce the activation of MCs, while the inactivated form of the drug (iCORM), unable to release CO, was ineffective. PMN significantly increased the production of ROS upon activation with fMLP and, consistently with the hypothesis that superoxide anion plays a role in MC activation, the treatment of the cells with SOD (300 IU/ml) mimicked the effects of CORM-3. Finally, CORM-3 also reduced the activation of human PMN, assessed as the membrane expression of CD11b (evaluated through flow cytometry). The inactivated form of CORM-3 and SOD were ineffective.
In conclusion, CORM-3 was effective in reducing fMLP-PMN-induced MC activation. The effect was mediated by the release of CO, since the iCORM was ineffective. We can also suggest an involvement of superoxide anion, since the activation of MCs was reverted incubating the cells with SOD.
30
Prevention of Antigen-Induced Bronchoconstriction by Epigallo-catechine-3-Gallate in Actively Sensitized Guinea Pigs
Lucia Giannini1, Caterina Uliva1, Daniele Bani2, Rosanna Mastroianni1, Ylenia Suzuki3, Silvia Nistri2, Pier Francesco Mannaioni1, Hisanori Suzuki4, Emanuela Masini1
1Department of Preclinical and Clinical Pharmacology, 2Department of Anatomy, Histology and
Forensic Medicine, University of Florence, 50139 Florence, 3Pediatric Clinic and Neurology,
University of Verona, 37134 Verona, Italy
It has been showed that nitric oxide (NO) plays a role in asthma, even if the pathophysiological connection between NO and asthma remains uncertain. In this study we used an animal model of asthma-like reaction, to provide insight into the possible role of NOS-derived NO in the pathophysiology of early asthma and to test the possible therapeutic effect of epigallocatechin-3-gallate (EGCG), a polyphenol present in green tea that enhances NO synthase (NOS) activity. For comparison, we used epicatechine (EC), which shares antioxidant but not NOS-modulating properties with EGCG.
Ovalbumin-sensitised guinea pigs placed in a respiratory chamber were challenged with ovalbumin. EGCG (25 mg/kg b.wt.) or EC (25 mg/kg b.wt.) were given i.p. 30 min before ovalbumin challenge. We analysed latency time for the onset of respiratory abnormalities; cough severity; duration of dyspnoea; lung tissue histopathology; mast cell activation (by granule release); leukocyte/eosinophilic infiltration (by major basophilic protein and myeloperoxidase); oxygen free radical-mediated injury (by nitrotyrosine and 8-hydroxy-2-deoxyguanosine); NOS activity; bronchial inflammatory response (by TNF-α in bronchoalveolar lavage, BAL).
Severe respiratory abnormalities appeared in the sensitised animals soon after the antigen challenge, accompanied by bronchoconstriction, alveolar inflation and a marked increase in the assayed parameters of inflammatory cell recruitment, free radical lung injury and release of proinflammatory molecules in BAL fluid. This was associated with marked depression of constitutive NOS activity. Pretreatment with EGCG, but not EC, significantly reduced all the above parameters and sustained endothelial-type NOS activity.
These findings indicate that EGCG, probably by modulating NOS activity, can counteract allergic asthma-like reactions in sensitised guinea pigs and suggest that it may be useful for the treatment of asthma in the future.
31 in Histidine Decarboxylase Knockout Mice
Agnes Koncz1, 2, Gyorgy Nagy1, 3, Mercedesz Mazan1, Edit Buzás1, Andras Falus1
1Semmelweis University, Department of Genetics, Cell- and Immunobiology, Budapest; 2National Medical Center, Inst. Hematology and Immunology, Budapest; 3Polyclinic, Hospitaller
Brothers of St. John of God, Budapest, Hungary
Histamine is a key regulator of the immune system, increased histamine production is a well known feature of many allergic diseases. Several lines of evidence suggest the role of histamine in T cell activation: T cells express both type 1 and type 2 histamine receptors, histamine inhibits the production of Th1 cytokines such as IL-2 and interferon-γ (IFN) and enhances the secretion of Th2 cytokines. IFN-γ was previously shown to regulate nitric oxide (NO) production. NO is a multifunctional intracellular and intercellular messenger, NO mediates lymphocyte proliferation differentiation and apoptosis. Our previous work indicates that NO is necessary for effective T cell activation.
To study the role of histamine in T cell activation, we investigated T cell signal transduction in histidine decarboxilase knockout (HDC-KO) and wild type mice. Splenocytes from wild type and HDC-KO mice were isolated following in vivo stimulation with complete Freund’s adjuvant. NO, cytoplasmic Ca2+ concentrations and reactive oxygen intermedier (ROI) levels were determined by flow cytomerty. IFN-γ mRNA level was measured by RT PCR.
Increased INF-γ mRNA level of HDC-KO splenocytes was associated with a markedly increased (2,5 fold, p=0,00086) NO production, compared to splenocytes of wild type animals. Activation of T cells through the TCR initiates a biphasic elevation in cytosolic Ca2+ concentration, a rapid initial peak observed within seconds and a plateau phase lasting up to 48 h. In response to Con-A stimulation, rapid Ca2+ fluxing was diminished (p<0.05) while the plateau phase was increased (p=0.0024) in KO T cells. T cell activation-induced NO and superoxide signals were similar in HDC-KO and wild type mice (p=0.45; p=0.23 respectively).
32
Histamine Influences the DNA Interaction of STATs in Human Lymphocytes
Friedhelm Diel, Hannelore Borck, Helena Meng
Institut für Umwelt und Gesundheit (IUG) and University of Applied Sciences, FB Oe, Biochemistry, Marquardstrasse 35, D-36039 Fulda, Germany
As signal transduction in thymocytes is correlated to signal transducer and activator of transcription (STAT), and histamine is an important mediator in allergic diseases the question arises whether histamine acting through histamine receptors (HR) induces crosstalk or modulation of the JAK/STAT pathway – especially the STAT1 interaction at the nuclear level.
Phytohemagglutinin (PHA) stimulated PBMC from atopic (n = 4) and non-atopic (n = 3) donors which served as an ex vivo model for measurements of the effect of histamine, clobenpropit (H3R antagonist, H4R agonist), thioperamide (H3R/H4R antagonists) and the neutral H4R antagonist JNJ7777120 on STAT1. DNA-binding was examined by electrophoretic mobility shift assay (EMSA) technique using STAT1 oligonucleotide (5´- CATGTTATGCATATTCCTGTAAGTGAAAA-3´; Metabion, Martinsried, Germany).
In agreement with previous results IL-4 production was increased and IFN-γ was decreased in the atopic as compared to the non-atopic group at the end of the lymphocyte 3-day culture period. This was correlated to down-regulation of STAT1 signalling in atopic individuals. Western blots showed STAT1α (91kDa) and additionally an 118kDa-band from which a 28kDa-fragment was cleaved in both groups.
The addition of histamine or clobenpropit led to a significant suppression of the formation of STAT1α in the non-atopic group. The STAT1α production in this group was enhanced by the H4R antagonists (thioperamide or JNJ7777120). The phosphorylation of STAT1 was also regulated by the H4R, as reflected by EMSA experiments showing decreased DNA-binding in the atopic group. Interestingly, the 28kDa-STAT1 DNA-binding was significantly suppressed by histamine (p<0.05), but significantly enhanced by the neutral H4-antagonist JNJ7777120 in both groups (p<0.001).
These results suggest that histamine influenced STAT1 downstream and modulated promoter gene interaction of the 28kDa-STAT1-fragment.
33 Miguel A Medina, José F Aldana, Francisco R Villatoro, Gonzalo Claros, José Luis Urdiales, Oswaldo Trelles, Francisca Sánchez-Jiménez
Departments of Molecular Biology and Biochemistry, Computer Architecture and Computational Languages. University of Málaga, Málaga, E-29071, Spain
Histamine, polyamines and other biogenic amines have pleiotropic physiological effects and the impairment of their metabolism is associated with multiple highly prevalent pathological situations [1]. New systemic approaches are needed for a more efficient advance of knowledge in order to control biogenic amine metabolism and its implication in pathophysiological conditions [2].
To achieve this goal, we use a practical approach: to generate an interconnected database to obtain emergent information on the molecular basis of biogenic amine metabolism and functions. We will build a bilingual (English-Spanish) online platform containing three working areas: an ontology-based integration area for the available information on the components of the system, a predictive and a training area. With respect to the predictive area, we are working on: i) protein modelling, that has allowed us to obtain the first 3D structural model for mammalian histidine decarboxylase [3]; ii) we have also built the first mathematical model integrating polyamine, arginine and sulfur amino acid metabolism (unpublished results). The architecture of the project is available in its provisional web page (http://av.bmbq.uma.es/asp). The results of this pilot project will be opened to students and scientists from other research centres and countries.
In conclusion, our project is expected to contribute to advance in the systemic integration of research on histamine, polyamine and other biogenic amines. European groups working on fully characterized experimental models are strongly encouraged to join us.
[1] Medina et al. Critic Rev Biochem Mol Biol 2003;38:23 [2] Medina et al. J Cell Mol Med 2005;9:854
[3] Moya-García et al () BioEssays 2005;27:57
Supported by National Institute of Bioinformatics (Genoma-España Foundation), SAF2005-1812 (MEC), and REMA G03/007 (FIS, MSC), and PAI (Andalusian Government).
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Effects of Histamine on Triglyceride Breakdown in Mouse Adipocytes Christian Carpéné, Sandy Bour, Danielle Prévot, Carine Duffaut, Philippe Valet U586 INSERM, IFR 31, Bat. L3, CHU Rangueil, 31432 Toulouse, France
Recent studies led to consider white adipose tissue (WAT) as a potential target for diverse histamine actions: treatments with H3-antagonists lower body weight gain (Hancock & Brune. Expert Opin Investig Drugs 2005;14:223-41) while histamine-deficient mice exhibit increased adiposity. However, direct histamine effects on WAT are scarcely documented. Several reports have demonstrated that histamine stimulates lipolysis in dog and rat adipocytes via H2-receptor activation. Other studies have shown that high doses of histamine can induce lipolysis inhibition, as a consequence of amine oxidation by the amine oxidases (AOs) present in adipocytes and subsequent hydrogen peroxide production. The aim of this work was therefore to study the influence of chronic AO blockade on histamine metabolic effects, and to search whether histamine action on lipolysis is altered in obesity. Semicarbazide was administered during 8 weeks in the drinking water of mice in order to totally block copper-containing AOs. Mice rendered obese by high fat diet were also studied, together with their lean controls. Lipolytic responses were studied in adipocytes from intra-abdominal WAT while subcutaneous (sc)WAT served for determination of AO activity. Chronic semicarbazide treatment (100 mg/kg) totally blocked the semicarbazide-sensitive amine oxidase activity (SSAO) in WAT without changing monoamine oxidase (MAO) activity. Treatment lowered body weight gain. Histamine did not activate lipolysis in control adipocytes but was moderately lipolytic in semicarbazide-treated mice. On the contrary, body and WAT weights were increased in diet-induced obese mice while SSAO showed only a slight tendency to be increased in WAT. Histamine (10-100 µM) was unable to stimulate lipolysis in both control and obese mice. These studies demonstrate that, at least in rodents, changes in histamine metabolism can modify histamine action on lipolysis. However, AO expression in adipocytes is not altered enough with diet-induced obesity to influence histamine metabolism and direct control of fat cell lipolysis.
35 Erna Pap, Éva Pállinger, András Falus
Department of Genetics, Cell – and Immunobiology, Semmelweis University, Budapest, 1089 - Hungary
Histamine seems to have an indisputable role in reproductive functions.
In males histamine influences fertility: gonadal development, spermatogenesis and sexual behavior. In females the role of histamine in the reproductive processes has been described from several aspects. It is required for normal ovulation, for the process of menstrual cycle, for placental blood flow regulation. It regulates the contractile activity of the uterus and lactation as well. Histamine has multiple functions in the process of pregnancy due to its vasoactive, differentiating and growth-promoting characteristics. Histamine is produced either by mast cells and/or by different cells of the reproductive organs. Pre – and postimplantation events are accompanied by high histidine decarboxylase (HDC) enzyme activity. H1, H2 receptors and diamine-oxidase are co-expressed in both decidual and placental cells in humans. Furthermore, the expression of HDC is much higher in the placenta than in any other organ.
Despite these results, exploring the influence of histamine in reproduction has not been a highlighted field. To fill this gap, our group has studied the regulatory role of histamine in reproduction using the HDC knockout in vivo mouse model. We found differences in the androgen production, gonadal development and sexual behavior in males. We measured alterations in the length of the menstrual cycle in histamine deficient mice. We found histamine dependency and differences in placental functions such as steroidogenic enzyme and cytokine expression. We described the importance of histamine in the establishment of proper Th1/Th2/Th3 balance at the placental-maternal interface during pregnancy.
Hereby we intend to present an overall summarizing picture about the influence of histamine in reproduction, underlining its special importance during pregnancy.
